Enzyme-Linked Immunosorbent Assay (ELISA) is a widely used analytical technique in equine research for detecting and quantifying specific proteins, hormones, and antibodies in horse biological samples. This method relies on antigen-antibody interactions and employs enzyme-linked antibodies to produce a measurable signal, typically a color change, indicating the presence and concentration of the target molecule. ELISA is applicable in various areas of equine health, including the diagnosis of infectious diseases, monitoring of immune responses, and assessment of physiological conditions. It is valued for its specificity, sensitivity, and ability to process multiple samples simultaneously. This page compiles peer-reviewed research studies and scholarly articles that explore the applications, methodologies, and advancements of ELISA in equine science.
Kleiboeker SB, Loiacono CM, Rottinghaus A, Pue HL, Johnson GC.The North American West Nile virus (WNV) epizootic, which began in 1999, has caused significant morbidity and mortality in horses. Because experimental infection has failed to consistently produce encephalitis in inoculated horses, investigation of naturally occurring cases was used to optimize strategies for diagnosis of this disease. Although WNV RNA could be detected by reverse transcriptase-polymerase chain reaction (RT-PCR) performed on whole blood collected from both clinically affected horses and unaffected herdmates, the diagnostic sensitivity of this approach was low compared with IgM...
Kollias-Baker C, Maxwell L, Stanley S, Boone T.Cocaine is a naturally occurring alkaloid that is commonly abused by human-beings for its psychostimulatory effects. Occasionally, very small concentrations (i.e. <100 ng/mL) of the primary cocaine metabolite, benzoylecgonine (BZE) have been detected in urine collected from horses competing in athletic events. In this study urine samples, collected from four horses following the administration of 2.5 and 20 mg of cocaine sublingually and 50 mg of cocaine intravenously, were analyzed for the presence of cocaine and/or its metabolites by enzyme-linked immunosorbent assay (ELISA) and gas chrom...
Johnson A, Smith R, Saxne T, Hickery M, Heinegård D.Previous experiments have shown that addition of fragmented fibronectin can induce cartilage chondrolysis. In this study we investigated the fate of the collagen- and cell-binding molecules Cartilage oligomeric matrix protein (COMP) and chondroadherin. Methods: Equine articular cartilage explants were stimulated with the C-terminal and the N-terminal heparin-binding fragments of fibronectin respectively, and the conditioned media were analysed by both quantitative (ELISA) and qualitative (mass spectrometry, Western blots) methods. Results: Both COMP and chondroadherin were released in a dose-d...
Tamaki Y, Hirata H, Takabatake N, Bork S, Yokoyama N, Xuan X, Fujisaki K, Igarashi I.A Babesia caballi gene encoding the 134-kDa (BC134) protein was immunoscreened with B. caballi-infected horse serum. An enzyme-linked immunosorbent assay (ELISA) using recombinant BC134 protein could effectively differentiate B. caballi-infected horse sera from Babesia equi-infected or noninfected control horse sera. These results suggest that the recombinant BC134 protein is a potential diagnostic antigen in the detection of B. caballi infection.
Huang X, Xuan X, Xu L, Zhang S, Yokoyama N, Suzuki N, Igarashi I.An immunochromatographic test (BeICT) for the rapid detection of antibodies against Babesia equi was developed. It clearly differentiated B. equi-infected horses from B. caballi-infected and uninfected horses. The agreement with enzyme-linked immunosorbent assay results was 96.7% in the detection of field sera. The results suggest that BeICT is rapid, simple, reliable, and suitable for use to detect B. equi infection in the field.
Ollivier FJ, Brooks DE, Schultz GS, Blalock TD, Andrew SE, Komaromy AM, Cutler TJ, Lassaline ME, Kallberg ME, Van Setten GB.Healing of corneal ulcers in horses is often associated with profound corneal stromal fibrosis and scar formation resulting in visual impairment. Connective tissue growth factor (CTGF) is a fibrogenic cytokine involved in wound healing and scarring. The purpose of this study was to determine whether CTGF was present in the tear fluid of normal horse eyes and the eyes of horses with corneal ulcers in order to evaluate the role of CTGF in corneal wound healing and corneal scar formation. Methods: Tear fluid samples were collected from 65 eyes of 44 horses; 32 samples from normal eyes, 21 samples...
Studdert MJ, Hartley CA, Dynon K, Sandy JR, Slocombe RF, Charles JA, Milne ME, Clarke AF, El-Hage C.Five of 10 pregnant, lactating mares, each with a foal at foot, developed neurological disease. Three of them became recumbent, developed complications and were euthanased; of the two that survived, one aborted an equine herpesvirus type 1 (EHV-1)-positive fetus 68 days after the first signs were observed in the index case and the other gave birth to a healthy foal on day 283 but remained ataxic and incontinent. The diagnosis of EHV-1 myeloencephalitis was supported by postmortem findings, PCR identification of the virus and by serological tests with an EHV-1-specific ELISA. At the time of the...
Xu Y, Zhang S, Huang X, Bayin C, Xuan X, Igarashi I, Fujisaki K, Kabeya H, Maruyama S, Mikami T.The prevalence of equine piroplasmosis caused by Babesia equi and Babesia caballi in northeast China has remained unknown, although the People's Republic of China is recognized as an endemic country for the diseases. In the present study, we investigated the prevalence of equine piroplasmosis in Jilin province, a part of northeast China. A total of 111 serum samples were taken from horses in eastern Jilin, and examined for diagnosis of B. equi and B. caballi infections by the enzyme-linked immunosorbent assays with recombinant antigens, equi merozoite antigen-1 and P48, respectively. Of the 11...
Hodgkinson JE, Lichtenfels JR, Mair TS, Cripps P, Freeman KL, Ramsey YH, Love S, Matthews JB.We report the use of six oligoprobes designed from intergenic spacer region sequences to identify fourth-stage larvae (L4) of the tribe Cyathostominae. Oligoprobes were designed for identification of the following species: Cylicocyclus ashworthi, Cylicocyclus nassatus, Cylicocyclus insigne, Cyathostomum catinatum, Cylicostephanus goldi, and Cylicostephanus longibursatus. A seventh probe was designed as a positive control to identify all these members of the Cyathostominae. The intergenic spacer region was amplified by PCR using conserved primers. Initially, three oligoprobes were used in South...
Kweon CH, Kwon BJ, Ko YJ, Kenichi S.VP7, the sero-group common antigen, of African horsesickness virus (AHSV-4) was expressed in insect cells by recombinant baculovirus. To develop a specific diagnostic method, monoclonal antibody (Mab) against VP7 was prepared and investigated as diagnostic reagent with the baculovirus expressed VP7. However, the Mab against VP7 of AHSV cross-reacted with Chuzan virus by the indirect immunofluorescence assay (IFA), confirming the presence of conserved domain of VP7 among Orbiviruses. This study describes two types of ELISA; Mab linked indirect (I-ELISA) and competitive-ELISA (C-ELISA) using bac...
Blitvich BJ, Fernandez-Salas I, Contreras-Cordero JF, Marlenee NL, Gonzalez-Rojas JI, Komar N, Gubler DJ, Calisher CH, Beaty BJ.Serum samples were obtained from 24 horses in the State of Coahuila, Mexico, in December 2002. Antibodies to West Nile virus were detected by epitope-blocking enzyme-linked immunosorbent assay and confirmed by plaque reduction neutralization test in 15 (62.5%) horses. We report the first West Nile virus activity in northern Mexico.
Clausen PH, Chuluun S, Sodnomdarjaa R, Greiner M, Noeckler K, Staak C, Zessin KH, Schein E.From May to July 2000, a cross-sectional study was conducted to estimate the prevalence of Trypanosoma equiperdum in the horse population of the central province (Tuv aimag) of Mongolia. On average, four herds were selected from each of the 29 aimag subdivisions (119 herds). From each herd, 10 horses were sampled in proportion to sex and age categories in the respective herds (1190 horses). Sera from 1122 horses were analysed for T. equiperdum antibodies using two serological assays, the complement fixation test (CFT) and the enzyme-linked immunosorbent assay (ELISA). The crude estimate of the...
Blitvich BJ, Bowen RA, Marlenee NL, Hall RA, Bunning ML, Beaty BJ.We evaluated the ability of epitope-blocking enzyme-linked immunosorbent assays (ELISAs) to detect West Nile virus (WNV) antibodies in domestic mammals. Sera were collected from experimentally infected horses, cats, and pigs at regular intervals and screened in ELISAs and plaque reduction neutralization tests. The diagnostic efficacies of these techniques were similar.
Kalina WV, Pettigrew HD, Gershwin LJ.Equine disease with an allergic etiology is common. Environmental antigens most often implicated as allergens in horses include molds, dusty hay, grass pollen, hay dust mites, and insect saliva. Although intradermal testing with allergen is a useful diagnostic tool for some species, skin testing frequently produces false positive results in horses. Allergen deprivation as a diagnostic tool is often impossible and at best it is ineffective at diagnosing the specific allergic reactivity. Synthesis of IgE after exposure to allergen is the instigator of the allergic process. While IgE exerts its e...
Illera JC, Silván G, Munro CJ, Lorenzo PL, Illera MJ, Liu IK, Illera M.An amplified enzymeimmunoassay (EIA) was validated for androstenedione in the serum of male horses. We will use the assay as a tool for the diagnosis of equine cryptorchidism. We will compare androstenedione EIA to the currently used methods (testosterone and estrone sulphate determinations). The study was conducted on 115 horses of pure Spanish and Arabian breeds, that included 30 geldings, 60 bilateral cryptorchids and 25 stallions. Androstenedione standard curve covered a range between 0 and 1 ng per well. Low detection limit was 1.54 pg/ml. Intra- and inter-assay coefficients of variation ...
Wagner HM, Balasuriya UB, James MacLachlan N.In an effort to further characterize the humoral immune response of horses to equine arteritis virus (EAV), direct and competitive enzyme-linked immunosorbent assays (c-ELISAs) were developed using monoclonal and polyclonal anti-sera to structural (G(L), N and M) and non-structural (nsp1) viral proteins. A nsp1-specific monoclonal antibody was produced to facilitate development of a c-ELISA to this protein. Data obtained using the various c-ELISAs confirm that the M protein is a major target of the antibody response of horses to EAV. However, none of the c-ELISAs that were developed were as se...
Wang T, Magnarelli LA, Anderson JF, Gould LH, Bushmich SL, Wong SJ, Fikrig E.Recombinant West Nile virus envelope (E) protein was examined in enzyme-linked immunosorbent assay (ELISA) to detect antibodies elicited during West Nile virus infection. Horses (nine of 10) and humans (six of six) with confirmed West Nile virus infection had IgG and/or IgM antibodies to the E protein. Antibodies to the recombinant West Nile virus membrane and nonstructural 1 proteins were not detected in any of these sera. An E protein-based ELISA may aid in the serological diagnosis of West Nile virus infection.
Kumba FF, Claasen B, Petrus P.A 15-year record of the results of horse sera from the Khomas region of Namibia tested by the complement fixation test for dourine at the Central Veterinary Laboratory in Windhoek before clearing the respective animals for export and competitive sport were subjected to statistical analysis. The range of percentage positive, taken as the apparent prevalence of dourine for the region, during the period of study, was 0-29.09%; the average regional level of apparent prevalence was 8.33%. These figures were thought to be lower than the real situation due to some bias in the sampling criteria. For m...
Giguère S, Hernandez J, Gaskin J, Prescott JF, Takai S, Miller C.The performance of four enzyme-linked immunosorbent assays (ELISAs) (ELISA-6939, ELISA-33701, ELISA-VapA, and ELISA-California) and an agar gel immunodiffusion test for diagnosis of Rhodococcus equi pneumonia in foals was evaluated. Antibody concentrations of foals with culture-confirmed R. equi pneumonia (n = 41) were compared to those of age-matched pasturemates that remained clinically healthy during the entire breeding season (n = 24). For each serological assay evaluated, selection of a low cutoff resulted in high sensitivity but low specificity. Increasing the cutoff value resulted in be...
Huang X, Xuan X, Yokoyama N, Xu L, Suzuki H, Sugimoto C, Nagasawa H, Fujisaki K, Igarashi I.The gene encoding a truncated merozoite antigen-2 (EMA-2t) of Babesia equi was cloned and highly expressed in Escherichia coli as a glutathione S-transferase fusion protein (G-rEMA-2t). Both G-rEMA-2t and rEMA-2t (after the removal of glutathione S-transferase) had good antigenicity. Either Western blot analysis with rEMA-2t or enzyme-linked immunosorbent assay (ELISA) with G-rEMA-2t clearly discriminated the sera of horses experimentally infected with B. equi from sera of horses infected with Babesia caballi and healthy horses, although rEMA-2t was not suitable for ELISA, probably owing to it...
Cuteri V, Takai S, Moscati L, Battistacci L, Pieramati C, Valente C.A serological survey of Rhodococcus equi infection was carried out on 602 blood samples collected from foals in central Italy. The assay was performed with an ELISA test using two different antigens prepared with reference strains of R. equi, ATCC 33071 and ATCC 6939. A positive reaction was obtained on 81 serum samples (13.45%) (OD > or = 0.3) using antigen ATCC 33071, and on 73 serum samples (12.12%) using antigen ATCC 6939. Although the frequency of the disease was not high, the serological positivity was about 13%. There was no statistically significant difference between males and fema...
Kumar S, Kumar Y, Malhotra DV, Dhar S, Nichani AK.Serial dilution and single dilution enzyme linked immunosorbent assays (ELISA) were standardised and their sensitivity and specificity were compared for serodiagnosis of Babesia equi infection. The antibody titres of 24 donkey sera of known identity were determined separately by serial dilution ELISA using three different B. equi antigens namely whole merozoite (WM), cell membrane (CM) and high speed supernatant (HSS). The ratios of the optical density (OD) of known positive and known negative sera at different serum dilutions were calculated and termed as the positive/negative (P/N) ratio. Th...
Hu WG, Alvi AZ, Fulton RE, Suresh MR, Nagata LP.A recombinant gene encoding a single-chain variable fragment (scFv) antibody against Venezuelan equine encephalitis virus (VEE) was cloned into a prokaryotic T7 RNA polymerase-regulated expression vector. A streptavidin-binding peptide gene fused to a 6His tag was attached downstream to the scFv gene. The recombinant fusion protein was expressed in bacteria as inclusion bodies that were subsequently solubilized with 8 M urea and renatured by an arginine system. Purification of the fusion protein was achieved by immobilized metal affinity chromatography. Enzyme-linked immunosorbent assay (ELISA...
Hirata H, Xuan X, Yokoyama N, Nishikawa Y, Fujisaki K, Suzuki N, Igarashi I.The efficacy of the Be82 gene product fused with glutathione S-transferase (GST/Be82) in an enzyme-linked immunosorbent assay (ELISA) for the diagnosis of Babesia equi infection was reported previously (H. Hirata et al., J. Clin. Microbiol. 40:1470-1474, 2002). However, the ELISA with the GST/Be82 antigen cross-reacted with Babesia caballi-infected horse sera, despite the high rate of detection of B. equi. These results suggested that GST/Be82 has an antigen in common with B. caballi or antigenicity similar to that of B. caballi. In the present study, we constructed a series of five clones wit...
The Journal of parasitologyJanuary 23, 2003
Volume 88, Issue 6 1239-1246 doi: 10.1645/0022-3395(2002)088[1239:QEOSTF]2.0.CO;2
Packham AE, Conrad PA, Wilson WD, Jeanes LV, Sverlow KW, Gardner IA, Daft BM, Marsh AE, Blagburn BL, Ferraro GL, Barr BC.Neospora hughesi is a newly recognized protozoan pathogen in horses that causes a myeloencephalitis similar to Sarcocystis neurona. There are no validated serologic tests using the gold standard sera that are currently available to detect specific N. hughesi antibodies and, thus, no tests available to detect antemortem exposure or estimate seroprevalence in the horse. The objectives of the present study were to establish a bank of gold standard equine sera through experimental infections with N. hughesi and to assess several serologic tests for the detection of related protozoan antibodies. Se...
Monzon CM, Mancebo OA, Russo AM.An ELISA test was used to determine the persistence of antibody levels in horses following treatment for Trypanosoma evansi. In 17 horses with T. evansi from two farms treated and cured with quinapyramine sulphate, ELISA antibody levels fell progressively post-treatment, but remained with positive results for 22.6 months in one horse, 12.8 months in a second, 4.1 months in another four and 2.3 months in three, whilst the rest became negative at 2.3 months. In two horses that suffered a post-treatment infection relapse the decrease in ELISA levels was only temporary, and a new increase in antib...
Hobo S, Yoshihara T, Oikawa M, Jones JH.An enzyme-linked immunosorbent assay (ELISA) was developed for equine surfactant proteins SP-A and SP-D in bronchoalveolar lavage fluid (BALF). Anti-equine SP-A or SP-D monoclonal antibodies (mAb) were produced by hybridoma technology, purified by the antibody purification reagent, and analysed by Western blotting analysis. The immunoreaction (two-site sandwich ELISA) with a mAb, peroxidase-labelled mAb and BALF sample was carried out simultaneously and analytical recovery and precision were assayed. Six mAb for SP-A and four mAb for SP-D were successfully cloned in limiting dilution to monocl...
Daly P, Doyle S.Equine herpesvirus-1 (EHV-1) infection is of significant animal welfare and economic importance. Yet, no standardised molecular techniques are available for diagnosis or confirmation of viral infection. The purpose of this study was to develop a standardised and quantitative assay system for the reliable detection of EHV-1 infection which was capable of eliminating the likelihood of false negative results. A region within the EHV-1 glycoprotein B gene was amplified by polymerase chain reaction (PCR), cloned and subjected to site-directed mutagenesis to generate a control plasmid, amplifiable b...
Eenoo PV, Delbeke FT.Salbutamol is a beta-adrenergic agonist that is used in the treatment of asthma in humans and chronic obstructive pulmonary disease in horses. Because of its stimulating and growth promoting properties, it is prohibited by horse racing authorities. Recently a number of adapters (eg Equinehaler) have been designed, allowing the use of metered dose inhalers (MDI) approved for human use. However, information on detection times of salbutamol after administration of salbutamol in therapeutic doses by inhalation is lacking. In this study, 2 mg salbutamol (Ventolin) was administered to four standardb...
Laverty S, Okouneff S, Ionescu M, Reiner A, Pidoux I, Webber C, Rossier Y, Billinghurst RC, Poole AR.Articular osteochondrosis (OCD) occurs in both man and animals. The etiology remains to be determined. Studies of OCD lesions in animals may provide clues as to its pathogenesis. The aim of our study was to determine whether there was evidence for increased degradation namely proteoglycan (PG) release and type II collagen cleavage in articular cartilage harvested from OCD lesions. We examined ex vivo explants at post-mortem from equine OCD lesions and macroscopically normal site and age matched cartilage. These were cultured over a 10 day period in serum-free medium. Type II collagen cleavage ...
Dehghan Rahimabadi P, Nazari A, Kamyabi M, Mosavari N.Glanders is the oldest and very contagious disease among horses caused by Burkholderia mallei. The disease occurs as a chronic form in horses. Hence, because of the prolonged shedding, numerous horses can potentially get infected by one horse with glanders. Glanders is endemic in Iran and this causes occasional occurrence in horse population of the country. The aim of this study was to determine the incidence of B.mallei infection in horses in two central provinces of Iran. A total of 517 serum samples were collected from stable horses in Tehran and Alborz provinces. Among the studied horses, ...
Favaro PF, Reischak D, Brandao PE, Villalobos EMC, Cunha EMS, Lara MCC, Benvenga GU, Dias RA, Mori E, Richtzenhain LJ.The equine influenza virus (EIV) H3N8 subtype is responsible for all EIV outbreaks worldwide while the H7N7 subtype is less pathogenic and is considered extinct as it has not been confirmed in outbreaks since 1980. Although EIV is enzootic in Brazil, few reports describe the actual EIV antibody status in the country. The aims of this study were: - to evaluate the efficiency of different serum treatments described by the World Organisation for Animal Health (OIE) and the World Health Organization (WHO) to remove non-specific haemagglutination inhibitors for the haemagglutination inhibition (HI)...
de la Rebière de Pouyade G, Franck T, Salciccia A, Deby-Dupont G, Grulke S, Heyden LV, Sandersen C, Serteyn D.Equine neutrophil elastase (NE) is a protease released in inflammatory diseases and participating in tissue destruction. To measure NE in horse plasma to assess its role in pathological conditions, we purified elastase from equine neutrophils by a double step chromatography and obtained a pure protein of 27 kDa, 4 kDa smaller than the NE 2A previously purified (Scudamore et al., 1993; Dagleish et al., 1999), which was likely to be NE 2B. We developed an ELISA by using two specific polyclonal antibodies obtained from rabbit and guinea pig. The sandwich complex was detected using a secondary ant...
Salud publica de MexicoJune 26, 2007
Volume 49, Issue 3 210-217 doi: 10.1590/s0036-36342007000300006
Fernández-Salas I, de Lourdes Garza-Rodríguez M, Beaty BJ, Jiménez JR, Rivas-Estilla AM.To investigate the presence of WNV in birds, horses and humans in northeast Mexico. Methods: Serum samples from 33 birds, 24 horses and 237 humans were screened by ELISA for Anti-WNV antibodies. Human serum samples were also screened for WNV RNA using an RT-PCR assay. Results: Positive sera were found in three birds and 15 horses. Forty percent of the human serum samples were positive for IgG antibodies and 0% for IgM antibodies and viral RNA. Conclusions: The results of this study show that WNV is present in northeast Mexico and it is a new emergent infectious agent that represents a challeng...
Aribam SD, Ogawa Y, Matsui H, Hirota J, Okamura M, Akiba M, Shimoji Y, Eguchi M.Serotyping is an important element for surveillance of Salmonella. In this study, an anti-O:4 Salmonella monoclonal antibody-based competitive enzyme-linked immunosorbent assay that could identify Salmonella infection in cow, pig, horse, and chicken was developed. This detection system can therefore be useful for a wide range of animals and for humans.
Kalina WV, Pettigrew HD, Gershwin LJ.Equine disease with an allergic etiology is common. Environmental antigens most often implicated as allergens in horses include molds, dusty hay, grass pollen, hay dust mites, and insect saliva. Although intradermal testing with allergen is a useful diagnostic tool for some species, skin testing frequently produces false positive results in horses. Allergen deprivation as a diagnostic tool is often impossible and at best it is ineffective at diagnosing the specific allergic reactivity. Synthesis of IgE after exposure to allergen is the instigator of the allergic process. While IgE exerts its e...
Winston S, Fiscus S, Hesterberg L, Matsushita T, Mildbrand M, Porter J, Teramoto Y.The development of three separate rapid ELISAs for detecting antibodies in host serum to three different viruses is described. These include: 1. A direct antigen assay using enzyme labelled anti-canine Ig for detecting antibodies to canine parvovirus, 2. A competitive ELISA using a feline infectious peritonitis virus-specific monoclonal antibody labelled with enzyme, and 3. A competitive ELISA using an equine infectious anemia virus-specific monoclonal antibody and enzyme labelled antigen, p. 26. The utility and benefits of each of the three approaches is emphasized.
Tamaki Y, Hirata H, Takabatake N, Bork S, Yokoyama N, Xuan X, Fujisaki K, Igarashi I.A Babesia caballi gene encoding the 134-kDa (BC134) protein was immunoscreened with B. caballi-infected horse serum. An enzyme-linked immunosorbent assay (ELISA) using recombinant BC134 protein could effectively differentiate B. caballi-infected horse sera from Babesia equi-infected or noninfected control horse sera. These results suggest that the recombinant BC134 protein is a potential diagnostic antigen in the detection of B. caballi infection.
Yapklç O, Yavru S, Kale M, Bulut O, Simşek A, Sahna KC.In this study, 162 horses, 80 donkeys and 51 mule serum samples were collected in Konya city. Additionally, 64 horse serum samples from Ankara and 49 samples from Kayseri city were included in the study. A total of 406 serum samples were examined by agar gel immunodiffusion (AGID) and enzyme-linked immunosorbent assay (ELISA) for antibody to equine infectious anaemia virus (EIAV) and no positive result was detected.
Elsheikha HM, Mansfield LS.Sarcocystis neurona is an important protozoal pathogen because it causes the serious neurological disease equine protozoal myeloencephalitis (EPM). The capacity of this organism to cause a wide spectrum of neurological signs in horses and the broad geographic distribution of observed cases in the Americas drive the need for sensitive, reliable and rapid typing methods to characterize strains. Various molecular methods have been developed and used to diagnose EPM due to S. neurona, to identify S. neurona isolates and to determine the heterogeneity and evolutionary relatedness within this specie...
Eule JC, Wagner B, Leibold W, Deegen E.We investigated 30 healthy eyes and 41 eyes with ERU from 57 horses. The total immunoglobulin titers and titers of IgGa, IgGb, IgM were measured in aqueous humour, vitreous and serum using different ELISA techniques. Every sample investigated contained detectable amounts of immunoglobulins. Compared to control eyes significantly increased titers were found in the aqueous humour and vitreous of the ERU eyes for all immunoglobulin isotypes studied (p < or = 0.01). While IgM was detected in only 2 out of thirty aqueous humour and in none of the thirty vitreous samples of healthy eyes, 79.6% of sa...
Heskett KA, Mackay RJ.To investigate the use of a specific antibody index (AI) that relates Sarcocystis neurona-specific IgG quotient (Q(SN)) to total IgG quotient (Q(IgG)) for the detection of the anti-S neurona antibody fraction of CNS origin in CSF samples obtained from horses after intragastric administration of S neurona sporocysts. Methods: 18 adult horses. Methods: 14 horses underwent intragastric inoculation (day 0) with S neurona sporocysts, and 4 horses remained unchallenged; blood and CSF samples were collected on days - 1 and 84. For purposes of another study, some challenged horses received intermitten...
Balkaya I, Simsek S.Cystic echinococcosis (CE), caused by hydatid cysts, is a widespread and hazardous disease in humans and animals worldwide. The disease is very common in Turkey, causing serious economic losses and public health problem. In this study, the seroprevalence of equine CE was determined by enzyme-linked immunosorbent assay (ELISA). Methods: Partially purified cyst fluid antigen from sheep hydatid cyst fluid was used as antigen in ELISA. A total of 250 equids consisting of 206 donkeys and 44 horses from various regions of Erzurum province of Eastern Turkey. Results: Anti- Echinococcus granulosus ant...
Cuteri V, Takai S, Moscati L, Battistacci L, Pieramati C, Valente C.A serological survey of Rhodococcus equi infection was carried out on 602 blood samples collected from foals in central Italy. The assay was performed with an ELISA test using two different antigens prepared with reference strains of R. equi, ATCC 33071 and ATCC 6939. A positive reaction was obtained on 81 serum samples (13.45%) (OD > or = 0.3) using antigen ATCC 33071, and on 73 serum samples (12.12%) using antigen ATCC 6939. Although the frequency of the disease was not high, the serological positivity was about 13%. There was no statistically significant difference between males and fema...
Victor S, Lampa E, Rask Andersen A, Gafvelin G, Grönlund H, Elfman L.Horse allergens are less studied than allergens from other furry animals and these allergens must be evaluated to understand the complexity of allergy to horses. The aims of this study were to develop assays for the horse allergens Equ c 1 and Equ c 2 in dander and saliva and to determine their levels in ten horse breeds. The study also included a comparison of these findings with previous results on the levels of Equ c 4 performed on the same study population. The study population included 170 horses from 10 horse breeds including American Curly and Russian Bashkir horse, which have been sugg...
Vanniasinkam T, Barton MD, Das TP, Heuzenroeder MW.This chapter describes a strategy for mapping linear B-cell epitopes of proteins using synthetic biotinylated peptides in an ELISA.A set of overlapping peptides were designed based upon a known amino acid sequence of the target protein, VapA (Virulence-associated Protein A) of the bacterium Rhodococcus equi, an important pulmonary pathogen in foals. The peptides synthesized as biotinylated peptides were coated directly onto micro titer plates which had been pre-coated with NeutrAvidin™ and used to screen sera from foals confirmed to have R. equi disease. A linear B-cell epitope was identifie...
Allen WR, Mathias S, Lennard SN, Greenwood RE.Rapid enzyme-based immunoassays were used to measure concentrations of oestradiol-17 beta and progesterone in daily blood samples recovered throughout oestrus and for a few days after ovulation from 34 Thoroughbred and 8 pony-type maiden, barren and foaling mares. The first detectable fall in oestradiol-17 beta levels occurred in 88% of the mares within the interval -72 to 0 h with respect to ovulation and in 65% of mares within the interval of -48 to 0 h. The results indicated that serial daily hormone assays of this type could, in a high proportion of animals, predict a correct time for a si...
Maeda K, Mizukoshi F, Hamano M, Kai K, Kondo T, Matsumura T.Recently, a novel 12-mer B-cell epitope, MKNNPIYSEGSL, in the type-specific region of equine herpesvirus 1 (EHV-1) glycoprotein G (gG) was identified and used as an antigen for enzyme-linked immunosorbent assay (Maeda et al., J. Clin. Microbiol. 42:1095-1098, 2004). Although our prototype strain, TH20p, possesses two repeat sequences containing the B-cell epitope, the EHV-4 NS80567 strain has two repeat sequences that are not identical. One repeat sequence stretch contained the B-cell epitope, while the other contained the 11-mer, MKNNPVYSESL (underlining indicates a different amino acid). In ...
Jakob HP, Eckert J, Jemmi T, Gottstein B.For many decades trichinellosis has not been reported among Swiss domestic pigs. Considering the fact that Trichinella occurs in a sylvatic cycle in Switzerland, a study was designed to reevaluate the present epidemiologic situation by investigating 10,904 fattening pigs, 218 pigs with free access to pasturage or being kept on an alp, 104 domestic boars, 106 horses, 44 wild boars and 538 foxes using a direct and an indirect diagnostic technique (digestion method and serology with ELISA and an excretory/secretory antigen, respectively). The digestion method was performed according to EC-guideli...
Ollivier FJ, Brooks DE, Schultz GS, Blalock TD, Andrew SE, Komaromy AM, Cutler TJ, Lassaline ME, Kallberg ME, Van Setten GB.Healing of corneal ulcers in horses is often associated with profound corneal stromal fibrosis and scar formation resulting in visual impairment. Connective tissue growth factor (CTGF) is a fibrogenic cytokine involved in wound healing and scarring. The purpose of this study was to determine whether CTGF was present in the tear fluid of normal horse eyes and the eyes of horses with corneal ulcers in order to evaluate the role of CTGF in corneal wound healing and corneal scar formation. Methods: Tear fluid samples were collected from 65 eyes of 44 horses; 32 samples from normal eyes, 21 samples...
Current drug deliveryJanuary 31, 2012
Volume 9, Issue 6 637-644 doi: 10.2174/156720112803529747
da Costa MH, Sant'Anna OA, Quintilio W, Schwendener RA, de Araujo PS.Liposomes have been used since the 1970's to encapsulate drugs envisaging enhancement in efficacy and therapeutic index, avoidance of side effects and increase in the encapsulated agent stability. The major problem when encapsulating snake venoms is the liposomal membrane instability caused by venom phospholipases. Here the results obtained encapsulating Crotalus durissimus terrificus and a pool of Bothropic venoms within liposomes (LC and LB, respectively) used to produce anti-venom sera are presented. The strategy was to modify the immunization protocol to enhance antibody production and to ...
Gong ZL, Liu GY, Xie JR, Chai HP, Zhang LY, Li ZX, Tian ZC, Wang L, Liu JG.To clone and express BC48 gene of Babesia caballi, and to establish an indirect ELISA for the diagnosis of B. caballi in equine animals. Methods: The genomic DNA of B. caballi was extracted from the infected donkey blood. BC48 gene was amplified by PCR. The PCR product was cloned into expression plasmid pET28a, and expressed in E. coli BL21 with IPTG induction. The recombinant protein was purified by Ni-NTA affinity chro-matography and was used as a diagnostic antigen to establish an indirect ELISA. The reaction conditions of the indirect ELISA were optimized. Specificity and sensitivity of th...
Ponthier J, Franck T, Parrilla-Hernandez S, Niesten A, de la Rebiere G, Serteyn D, Deleuze S.Myeloperoxidase (MPO) is a pro-oxidant enzyme associated with decreased motility in thawed equine semen. This study aimed to describe MPO concentration, activity and subunits in raw and thawed semen and to correlate these data with motilities in raw and thawed semen. Semen samples from five stallions were collected four times. Motilities were assessed in raw and thawed semen. MPO assays were performed in raw seminal plasma, raw sperm-rich pellet and thawed semen. Total and active MPO concentrations were, respectively, assayed by enzyme-linked immunosorbent assay and specific immunological extr...
Dzierzecka M, Kita J.The investigations aimed to establish the reliability of the chosen serological tests designed for the diagnosis of Lyme borreliosis in horses. The investigations were carried out in five Horse Breeding Centres (OHK). Statistical analysis methods were used to determine sample size for particular centres: Krasne (Kr)--49, Łack (Ł)--21, Walewice (W)--111, BogusŁawice (B)--17, Kozienice (K)--61. The experimental material comprised the chosen horses from which blood samples were collected in order to obtain sera. The test used for indirect immunofluorescence assay (IFA No 75941, Bio-Mérieux) i...
Adamu SG, Hassan M, Ardo MB.A cross-sectional survey was conducted to determine the seroprevalence and risk factors influencing the presence of Brucella spp. antibodies in donkeys in Yobe south senatorial zone, Nigeria. The study was aimed at determining the importance of Brucella spp. infection in donkeys (Equus asinus). A total of 200 sera samples from of 105 males and 95 female donkeys were collected and screened for brucellosis using the rose bengal plate test (RBPT) and the indirect enzyme-linked immunosorbent assay (iELISA). Data obtained were analyzed to determine associations and risk factors. The analysis reveal...
Morris DD, Whitlock RH, Corbeil LB.An equine antiserum to core lipopolysaccharide was produced by inoculation of 6 horses with a boiled cell bacterin made from the J-5 mutant of Escherichia coli O111:B4. The antiserum immunoglobulin G titer to J-5 mutant E coli, as determined by enzyme-linked immunosorbent assay, was 1:15,006. Pooled serum prepared before inoculation (preimmune serum) had a J-5 immunoglobulin G titer of 1:350. The J-5 antiserum was tested for its protective efficacy in sublethal endotoxemia in 14 horses. Four horses served as nontreated controls and were given nothing before endotoxin challenge exposure (10 mic...
Magnarelli LA, Bushmich SL, Anderson JF, Ledizet M, Koski RA.A polyvalent ELISA and plaque reduction neutralization tests (PRNTs) were used to measure serum antibodies to West Nile virus (WNV) in horses naturally exposed to or vaccinated against this flavivirus in Connecticut and New York State, USA. Relying on a PRNT as a 'gold standard', the main objective was to validate a modified ELISA containing a recombinant WNV envelope protein antigen. It was also important to assess specificity by testing sera from horses that had other, undiagnosed illnesses. Sera for the latter study were obtained from 43 privately owned horses during 1995-1996. Analyses by ...
Ueda Y, Sanai Y, Homma JY.Enzyme-linked immunosorbent assay (ELISA) was used for detection of immunoglobulin (Ig) M and IgG antibodies against a serologically common antigen (original endotoxin protein), protease, and elastase of Pseudomonas aeruginosa. The P aeruginosa antibody in horse sera was measured, using ELISA. Horseradish peroxidase-labeled rabbit anti-horse IgM and IgG antibodies were used for enzyme-labeled antibody conjugate. 5-Aminosalicylic acid and H2O2 were used for substrate. Sera collected from a vaccinated horse, a newborn foal, and 72 healthy racehorses were investigated for antibodies against P aer...
Secor EJ, Matychak MB, Felippe MJ.This study aimed to determine whether TNF-α is transferred to equine neonates via colostrum and the relationship between TNF-α and IgG concentrations in the equine neonate. Colostrum, presuckle and postsuckle foal serum samples were collected from healthy mares and their foals. Equine TNF-α ELISA and IgG SRID kits were used to determine the concentrations of TNF-α and IgG, respectively. Statistical analysis was performed using the Spearman rank correlation. TNF-α concentrations in all presuckle foal serum were below the limit of detection in 15/16 foals and increased in postsuckle foal se...
